Synthesis of Thiazole-2(3H)-thiones as Antimicrobial Agents Promoted by H3PW12O40-amino-functionalized CdFe12O19@SiO2 Nanocomposite

INTRODUCTION
Thiazoles show anticancer [1], antifungal [2], anti-inflammatory [3], anti-Candida [4], and antibacterial properties [5]. Some other examples of thiazoles like prominent drug molecules such as Pramipexole [6], Tiazofurin [7], Ritonavir [8], and Nizatidine [9] are applied for their high bioavailability, prolonged effects, and a slower onset (Scheme 1). These properties make thiazoles appealing goals in organic synthesis. Past reports on the synthesis of 1,3-thiazole derivatives have mentioned catalysts such as 1,8-diazabicyclo[5.4.0]undec-7-ene [10], perchloric acid-SiO2 [11], Bi(SCH2COOH)3 [12], triethylammonium hydrogen sulfate [13], and ytterbium(III) triflate [14]. Whereas these ways have substantial negative aspects including high reaction times, low yields, unwanted reaction conditions, and the use of costly and toxic catalysts. Appling environmental and green nanocatalysts, which can be quickly recycled at the end of reactions, has received great attention in recent years [15,16]. Nanocomposites have appeared as a proper group of heterogeneous catalysts due to their numerous usages in synthesis and catalysis [17-18]. Since these nanocomposites are often retrieved easily by simple workup, which prevents organic products from contamination, they may be investigated as  promising, safe, reusable, and greener catalysts compared to traditional ones [19-23]. Newly, magnetic nanocomposite particles (MNPs) have been utilized to immobilize organocatalysts, polymers, enzymes, and transition metal catalysts [24-26]. Different types of magnetic nanocatalysts have been improved to efficiently promote multi-component reactions for the synthesis of medicinal compounds [27-28]. These nanocomposites are recyclable and can be retrieved in consecutive trials without significantly decreasing their catalytic attributes [29-30]. The heteropolyacids are extensively applied as homogeneous and heterogeneous catalysts [31-32]. In this work, H3PW12O40–amino was tethered to magnetic nanocomposite particles for preparing a reusable magnetic catalyst. Furthermore, we considered a useful way for the synthesis of thiazole-2(3H)-thiones as antimicrobial agents by three-component reactions of CS2, 2-bromoacetophenone or 2-bromo-1-(4-methoxyphenyl)ethanone and a primary amine (Scheme 2).

EXPERIMENTAL SECTION
Chemicals and apparatus 
All materials were commercially purchased from Merck and Sigma-Aldrich. Powder X-ray diffraction (XRD) was registered on a Philips diffractometer of X’pert Company with monochromatized Cu Kα radiation (λ = 1.5406 Å). The microscopic morphology of the nanocatalyst was recorded by SEM (MIRA3). The thermogravimetric analysis (TGA) curves were investigated using a V5.1A Dupont 2000. The magnetic measurement of samples was studied in a vibrating sample magnetometer (VSM) (Kashan, Iran: Kavir Co). Fourier transform infrared measurements were conducted on Magna 550 instrument by using potassium bromide (KBr) plates. NMR spectra were recorded on a Bruker 400 MHz spectrometer with DMSO-d6 as the solvent and TMS as internal standard.

Preparation of CdFe12O19 nanoparticle:
The CdFe12O19 nanocomposites were synth-esized using sol-gel auto-combustion path of a solution of Cd(NO3)2.4H2O and Fe(NO3)3.9H2O with the molar ratio of 1 to 12 for Cd:Fe. Initially, cadmium nitrate tetrahydrate was solved in distilled water. Afterwards, pomegranate juice (15 mL) was added into the solution drop-wise under strong magnetic stirring for 30 min. Next, an aqueous solution containing Iron (III) nitrate nonahydrate was added to the solution and heated at 90 °C for 60 min to form a viscous gel. The gel was dried in an oven at 100 °C. The final residue was calcined at 800 °C for 120 min [33-34]. Meanwhile, the final remnants were powdered by mortar.

Preparation of CdFe12O19@SiO2 nanocomposites
The nano-CdFe12O19 powder (1g) was dispersed in 20 mL ethanol in ultrasonic bath and sonicated for 50 min at 40 ºC. A concentrated ammonia solution (1.5 mL) was added under magnetic stirring at 40 ºC for 30 min. Then, tetraethyl orthosilicate (TEOS, 1.0 mL) was added to the solution and stirred at 40 °C for 24 h. CdFe12O19@SiO2 nanocomposites were collected using an external magnet and washed with ethanol and dried in a vacuum for 24 h.

Preparation of amino-functionalized CdFe12O19@SiO2 nanocomposites
CdFe12O19@SiO2 (1 g) was added to the solution of 3-aminopropyltriethoxysilane (APTES) (2 mmol, 0.44 g) in dry toluene (20 mL) and refluxed for 20 h. Finally, amino-functionalized CdFe12O19@SiO2 nanocomposites were collected by a magnet, washed with double-distilled water and anhydrous ethanol, and dried at 80 ºC for 8 h.

Preparation of H3PW12O40-amino-functionalized CdFe12O19@SiO2 nanocomposites
Amino-functionalized CdFe12O19@SiO2 (1 g) was added to the solution of 0.3 g of H3PW12O40 in methanol (25 mL) and the reaction mixture was refluxed for 6 h. Finally, H3PW12O40-amino-functionalized CdFe12O19@SiO2 nanocomposites were collected by a magnet, rinsed with methanol, and dried at room temperature.

General procedure for preparing thiazole-2(3H)-thiones
A mixture of primary amine (1.0 mmol) and CS2 (1.0 mmol) in ethanol (8 mL) was stirred for 5 min and then 2-bromoacetophenone or 2-bromo-1-(4-methoxyphenyl)ethanone (1.0 mmol) and H3PW12O40-amino-functionalized CdFe12O19@SiO2 nanocomposites (4 mg) were added, and the mixture was stirred for the appropriate times, as determined by TLC (n-hexane/ethyl acetate 7:3). After the completion of the reaction, the nanocomposite was separated from the reaction before working up by an external magnet field. The solid rinsed with EtOH to get a pure product. All of the compounds were identified on the basis of their 1H NMR, 13C NMR, FT-IR, and elemental analyses, all of which were submitted in the supporting information.  
3-Benzyl-4-phenyl-1,3-thiazole-2(3H)-thione (4a): 
Colorless viscous oil; FT-IR (KBr): 3103, 3004, 1605, 1476, 1203 cm-1; 1H NMR (250 MHz, CDCl3): δ 4.91 (s, 2H, CH2), 6.04 (s, 1H, CH of alkene), 6.94–7.35 (m, 10H, CH, ArH). 13C NMR (62.5 MHz, CDCl3): δ 47.25, 98.85, 127.05, 127.45, 128.50, 128.55, 129.05, 133.30, 137.45, 154.85, 178.35, 197.20. Anal. Calcd. for C16H13NS2 (283): C, 67.81; H, 4.62; N, 4.94. Found: C, 67.71; H, 4.53; N, 4.74 %.

Determination of antimicrobial activity 
The antimicrobial activity of compounds is determined using Agar diffusion [35]. Streptomycin (10µg/well) as a standard drug was applied as a positive control for bacteria, and Nystatine (100 IU/well) was used for fungi; DMSO was used as a negative control. The results were considered for each tested compound as the average diameter of inhibition zones of bacterial and fungal around the wells in mm.

RESULTS AND DISCUSSION
The particle size and morphology of H3PW12O40-amino-functionalized CdFe12O19@SiO2 nanocomposites was considered by SEM which clearly revealed that the average size of the particles is about 80 nanometers (Figure 1). 
Figure 2 indicates the powder X-ray diffraction (XRD) pattern of H3PW12O40-amino-functionalized CdFe12O19@SiO2 nanocomposites. The pattern agrees correctly with the presented pattern for CdFe12O19 [33-34]. The crystallite size of the nanocatalyst calculated by the Debye–Scherer equation is about 84-86 nm.
Figure 3 shows the FT-IR spectra of CdFe12O19, CdFe12O19@SiO2, amino-functionalized CdFe12O19@SiO2 and H3PW12O40-amino-functionalized CdFe12O19@SiO2 nanocomposites. The FT-IR spectra of CdFe12O19 display the vibrations of the metal–oxygen stretching at 555-565 cm−1. The absorption peak at 3436 cm-1 is corresponded to the stretching vibrational absorptions of O-H. The bands at 467, 1078, 1636, and 3425 cm-1 are the characteristic absorptions of SiO2, which provides the evidence for the formation of a silica shell. The increase of the bands at 1572 cm-1 is a direct indication of the existence of the N-H bending vibration (Fig. 3c). The bands at 1080, 980, 880, and 773 cm−1, which are the fingerprint of the Keggin structure of H3PW12O40, are usually corresponded to P-O, W=O, W-O-W in corner shared octahedral, and W- O-W in edge shared octahedral [36-37] (Fig. 3d). 
The magnetic attributes of CdFe12O19 (b), CdFe12O19@SiO2 (c), amino-functionalized CdFe12O19 @SiO2 (d), and H3PW12O40-amino-functionalized CdFe12O19@SiO2 nanocomposites were measured with VSM (Figure 4). The amounts of saturation-magnetization for CdFe12O19 and H3PW12O40-amino-functionalized CdFe12O19@SiO2 nanocomposites are 12.80 and 1.98 emu/g, respectively. These results demonstrate that the magnetization properties are lessened using the coating. Furthermore, these results also indicate that the H3PW12O40 -amino-functionalized CdFe12O19@SiO2 nanocatalyst remains magnetic after coating. This is advantageous because magnetic nanocatalyst can be easily collected from the reaction media by an external magnet field over a short period of time.
Thermogravimetric analysis (TGA) peruses the thermal stability of the H3PW12O40-amino-functionalized CdFe12O19@SiO2 nanocomposites. The weight loss at temperatures below 200 ºC is due to the removal of physically adsorbed solvent and surface hydroxyl groups. The curve exhibited a weight loss of about 12 % from 220 to 600 ºC owing to the decomposition of the organic spacer grafting to the nanocomposite surface (Figure 5).
The energy-dispersive X-ray spectrum (EDS) of H3PW12O40-amino-functionalized CdFe12O19@SiO2 nanocomposites (Figure 6) displays that the elemental compositions are carbon, cadmium, oxygen, iron, tungsten, silicon, nitrogen, and phosphorus. 
We used the reaction of CS2, 2-bromoacetophenone and benzyl amine on 1 mmol scale as a model reaction and carried it out using CAN, NaHSO4, CdFe12O19, CdFe12O19@SiO2, amino-functionalized CdFe12O19 @SiO2 and H3PW12O40-amino-functionalized CdFe12O19@SiO2 nanocomposite.  We found that the reaction gave useful results using H3PW12O40-amino-functionalized CdFe12O19@SiO2 nanocomposite (4 mg) (Table 1). The best results were exemplified in Entry 11 with 94% yield. Further, we also reacted 2-bromoacetophenone or 2-bromo-1-(4-methoxyphenyl)ethanone with CS2 and other primary amines and uniformly found satisfactory results (Table 2, mean 86%).  The yield did not appear to be exclusively sensitive to the substituent groups.  The structures of the products were deduced from their 1H NMR, 13C NMR, and FT-IR. 
The reusability of H3PW12O40-amino-functionalized CdFe12O19@SiO2 nanocomposite (4 mg) was considered for the model reaction, and it was found that product yields lessened only to a very small extent on each reuse (run 1, 94%; run 2, 94%; run 3, 93%; run 4, 93%; run 5, 92%; run 6, 92%).  After the completion of the reaction, the nanocatalyst was separated by an external magnet. The catalyst was rinsed four times with ethanol and dried at room temperature for 10 h.
Scheme 3 displays a proposed mechanism for this reaction in the presence of H3PW12O40-amino-functionalized CdFe12O19@SiO2 nanocomposite as catalyst. Initially, the nucleophilic attack by amines on a carbon disulfide generates intermediate (I); the next step involves the nucleophilic attack of intermediate (I) on the methylene carbon of phenacyl bromide, leading to intermediate (II); then, ring closure was achieved by intramolecular attack of nitrogen at the carbonyl carbon to afford the 3-alkyl-4-phenyl-1,3- thiazole-2(3H)-thione derivatives. In this mechanism the surface atoms of H3PW12O40-amino-functionalized CdFe12O19@SiO2 nanocomposite activate the C=S and C=O groups for better reaction with nucleophiles. 
The antimicrobial activity of compounds is determined using Agar diffusion [35]. The results are exhibited in Table 3. The compounds 4b, 4e, 4f, and 4j have moderate growth inhibitory effects on Gram positive bacteria (Staphylococcus aureus, Bacillus subtilis; and Staphylococcus epidermidis). The compound 4b has moderate growth inhibitory effects on fungi.

CONCLUSIONS
In conclusion, we demonstrated an efficient way for the preparation of thiazole-2(3H)-thiones by three-component reactions of CS2, 2-bromoacetophenone or 2-bromo-1-(4-methoxyphenyl)ethenone, and a primary amine using H3PW12O40-amino-functionalized CdFe12O19@SiO2 nanocomposites in ethanol. The compounds 4b, 4e, 4f, and 4j have moderate growth inhibitory effects on Gram positive bacteria (Staphylococcus aureus, Bacillus subtilis; and Staphylococcus epidermidis). The compound 4b has moderate growth inhibitory effects on fungi. This protocol has a number of salient features including great yields in concise times, retrievability of the nanocatalyst, little nanocatalyst loading, and antibacterial activities for four compounds.